Assay and properties of N-acetylglucosamine 6-phosphate deacetylase from rat liver
N-Acetylglucosamine (GlcNAc) is generated, in liver and other tissues, as a product of the lysosomal degradation of complex carbohydrates. After diffusion into the cytosol, the monosaccharide is metabolized via catabolic or anabolic pathways, in which the first step is common to both pathways and consists of phosphorylation at C-6 by a specific GlcNAc kinase. When processed via the catabolic pathway, GlcNAc 6-phosphate is then deacetylated to glucosamine (GlcN) 6-phosphate. In the present study, they have developed a simple assay for the deacetylase catalyzing this reaction, in which /sup 3/H-acetate released from /sup 3/H-acetyl-labeled GlcNAc 6-phosphate is measured directly in a biphasic liquid scintillation counting system after acidification of the reaction mixture. The new assay has been used in partial purification of the deacetylase from rat liver and in analysis of kinetic and other properties of the enzyme. GlcN 6-phosphate and glucose 6-phosphate inhibited the deacetylase, while GlcNAc, GlcNAc 1-phosphate, GlcN 1-phosphate, and N-acetylgalactosamine had no effect. The activity of the enzyme was substantially inhibited by 5 mM Mn/sup 2 +/, Ni/sup 2 +/, or Cu/sup 2 +/, while Ca/sup 2 +/, Mg/sup 2 +/, and Ba/sup 2 +/ had essentially no effect at concentrations of 12.5 mM or lower. Co/sup 2 +/ stimulated at low concentrations (< 5 mM) but inhibited at higher concentrations. Gel chromatography on Sepharose CL-6B indicated that the enzyme had a mol. wt. greater than 160,000.
- Research Organization:
- Univ. of Alabama, Birmingham
- OSTI ID:
- 6060601
- Report Number(s):
- CONF-870644-; TRN: 87-037629
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 46:6; Conference: 78. annual meeting of the American Society of Biological Chemists conference, Philadelphia, PA, USA, 7 Jun 1987
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
BARIUM COMPOUNDS
BIOLOGICAL EFFECTS
CALCIUM COMPOUNDS
CARBOHYDRATES
METABOLISM
COBALT COMPOUNDS
COPPER COMPOUNDS
GLUCOSAMINE
BIOSYNTHESIS
MAGNESIUM COMPOUNDS
MANGANESE COMPOUNDS
NICKEL COMPOUNDS
OXIDOREDUCTASES
BIOASSAY
CHEMICAL PROPERTIES
FRACTIONATION
ACETATES
BIOLOGICAL PATHWAYS
CATIONS
CHROMATOGRAPHY
INHIBITION
LIVER
MOLECULAR WEIGHT
RATS
SCINTILLATION COUNTING
TRACER TECHNIQUES
TRITIUM COMPOUNDS
ALKALINE EARTH METAL COMPOUNDS
AMINES
ANIMALS
BODY
CARBOXYLIC ACID SALTS
CHARGED PARTICLES
COUNTING TECHNIQUES
DIGESTIVE SYSTEM
ENZYMES
GLANDS
HEXOSAMINES
HEXOSES
IONS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
MONOSACCHARIDES
ORGANIC COMPOUNDS
ORGANS
RODENTS
SACCHARIDES
SEPARATION PROCESSES
SYNTHESIS
TRANSITION ELEMENT COMPOUNDS
VERTEBRATES
560300* - Chemicals Metabolism & Toxicology
550201 - Biochemistry- Tracer Techniques